Effect of Disulfiram and Copper Plus Chemotherapy vs Chemotherapy Alone on Survival in Patients With Recurrent Glioblastoma

Key Points Question Does the addition of disulfiram and copper to chemotherapy improve survival for patients with recurrent glioblastoma? Findings In this randomized clinical trial of 88 patients with recurrent glioblastoma, the addition of disulfiram and copper to alkylating chemotherapy did not significantly improve survival at 6 months, as compared with alkylating chemotherapy only. Significantly more patients receiving disulfiram had adverse events of grade 3 or higher (34% vs 11%) and serious adverse events (41% vs 16%). Meaning These findings suggest that the addition of disulfiram and copper to alkylating chemotherapy should not be recommended for patients with recurrent glioblastoma.


Sample size
Assuming an improvement in proportion achieving 6-month survival from 60 % to 80 % we need a sample size of 128 in total (64 in each group; alpha 0.1, power 80 % and two sided test BACKGROUND Disulfiram (Antabus®) is a well-tolerated, cheap, generic drug that has been in use since the 1950s to treat alcoholism. There is now an increasing amount of independent preclinical data to support disulfiram as an anticancer agent. In glioblastomas add-on treatment to alkylating agents may offer additional benefit due to O 6 -methylguanine methyltransferase (MGMT, a DNA-repair enzyme) inhibition. The potency of disulfiram as an anticancer agent seems strengthened by copper. There is now anecdotal clinical evidence of disulfiram as an anticancer, for instance with improved survival in lung cancer patients. Also, disulfiram was positively associated with survival in a registry study. Collectively these results indicate a clinical meaningful response. So far no clinical studies have been published in glioma patients, but two small, uncontrolled studies are planned according to clinicaltrials.gov. with search 1 st November 2015.

OBJECTIVES
To investigate disulfiram and copper-supplement as add-on treatment in glioblastoma patients receiving alkylating chemotherapy. This will to our knowledge be the first planned RCT with disulfiram in glioblastoma patients, and will consequently serve as a proof-of concept study. PRIMARY OUTCOME and presenting with a first progression/recurrence documented by MRI. 2. Indication for treatment with chemotherapeutic alkylating agents (i.e. temozolomide OR lomustine (also known as CCNU) including the so called PCV treatment (procarbazine, CCNU,

Background
Gliomas are the most common malignant brain tumor, and they result in more years of life lost than any other tumor group. 1 The annual age adjusted incidence rate is approximately 9 per 100,000 in adults. 1 Glioblastoma, WHO grade IV astrocytoma, is the most common subtype and has a poor survival with median survival of only 10 months. 2 Extensive surgical resections followed by postoperative fractioned radiotherapy and concomitant and adjuvant temozolomide (an alkylating agent) prolong survival in glioblastoma (WHO IV) and is the mainstay of treatment. 3 Still, the median overall survival in the RCT by Stupp et al. was only 14.6 months. The effect of temozolomide seems at least in part dependent of O 6methylguanine methyltransferase methylation (MGMT, a DNA repair enzyme), leading to impaired DNA repair of damages caused by temozolomide within the tumor. 4 Unfortunately less than half of glioblastoma patients harbor MGMT methylation. 4 In the recurrent situation there is no true consensus on what to use. A common approach is to prescribe temozolomide again as long as the progression occurs more than 3-6 months after ended primary treatment, and especially if initial response was observed. However, in cases where progression is seen on ongoing or early after temozolomide treatment it is common practice to prescribe lomustine (also known as CCNU) 5 or PCV (procarbazine, CCNU, vincristine). Still, the treatment effect in this situation is modest and new treatments are clearly indicated.
Based on knowledge of pathophysiology and pharmacology, the 'repurposing' or 'repositioning' of approved drugs as treatments for other diseases is currently a topical issue relevant for industry, academia and governmental policies. According to Francis Collins, the Director of the National Institutes of Health, research in this direction should be 'an important focus of the NIH's proposed National Center for Advancing Translational Sciences'. 6 1.1. The case for Disulfiram (Antabus®)

Generic drug with well-known safety profile
Disulfiram is a cheap, generic drug that has been used since 1947 to treat alcoholism and is most commonly known as "Antabus®". 7 The drug is well tolerated (as long as one avoids alcohol), however some caution is necessary with respect to comorbidities and interactions. Health authorities (i.e. EMA and FDA) have already approved its use in humans, but only for the indication alcohol dependency.
Disulfiram safety is well documented. In alcohol dependent patients long-term use (> 1 year) has also been demonstrated to be safe and even normalization of liver enzymes initially elevated due to alcohol use has been observed during disulfiram treatment. 7,8 However, there are reports that in patients with alcohol dependence disulfiram may cause hepatitis in 1:30.000 cases. 7 There are also reported cases of neuropathy with a frequency of 1:15.000 treatment years, with typically dose-dependent occurrence with doses of > 500 mg/day and long-term treatment with the typical presentation after 1 year of treatment. 7 Optic neuritis may be more frequent in smokers, thus it may be worthwhile informing patients to quit before starting treatment (www.fass.se). However, neuropathy including optic neuritis is typically reversible if detected early. Psychosis and other acute confusional states beginning with fatigue and forgetfulness, rarely proceeding to ataxia or stupor has been reported, but psychiatric syndromes are common in alcohol dependent patients, and in a controlled study there were no differences in confusion/psychosis between groups receiving disulfiram or not. 7 Nevertheless, confusion appear to have occurred more often earlier when dosage regimens typically were > 500 mg/day. Because of the potentially fatal outcome of a disulfiram-alcohol reaction in a patient with heart disease disulfiram should normally not be offered patients with impaired cardiac function (i.e. heart failure). Still, the most important issue is total abstinence from alcohol. 7 In our context with cancer patients in need of chemotherapy, recent studies with disulfiram (or its metabolite) as add-on report minimal toxicities. 9, 10 The combination of chemotherapy and disulfiram in preexisting disulfiram users developing cancer is a common clinical practice. And a phase I study on disulfiram in glioblastoma patients taking temozolomide was well tolerated below 500 mg daily dose.

Preclinical evidence
Disulfiram has demonstrated a wide range of antineoplastic effects with potential effect in gliomas and this has been translated into promising effects in cell lines and cell cultures. [11][12][13][14][15][16][17] The anti-cancerous effect has also been demonstrated in different, independent animal models. 11,17 It is now suggested that disulfiram should be implemented in clinical trials 18 and indeed 2 small non-randomized studies are registered in clinicaltrials.gov when searching "glioma" AND "disulfiram" (www.clinicaltrials.gov, search date 01th November 2015). The drug may further potentiate alkylating damage possibly through inhibition of MGMT. 19

Clinical evidence to date
Dsiulfiram-copper complex (Cu-DSF) has been considered to be the (most) active antineoplastic metabolite of disulfiram in terms of cancer cells toxicity and ubiquitin proteasome system (UPS) interference.
In two small RCTs, one on breast cancer (64 patients) and one in metastatic lung cancer disulfiram (or one of its metabolites) has been tested with promising results on overall survival. 9,10 Also, in a small phase I study in glioblastoma promising results were observed. 20

Dose-rationale
The administration of treatment with disulfiram and copper is scheduled to start concomitant with alkylating chemotherapeutic treatment. The administration of disulfiram and copper supplement should be continued as adjuvant after chemotherapy withdrawal with continued administration as long as tumor targeted therapy is indicated.

Disulfiram:
Disulfiram as an anti-cancer agent has been suggested at a dose of 250 mg twice daily 18 although the pill in common in Norway and Sweden is 200 mg. A daily dose > 500 mg/day of disulfiram is associated with adverse-effects. Disulfiram is cheap, with a cost of administration of 400 mg a day below both 10 NOK and SEK.
Patients will take disulfiram (Antabus ® ) as a once daily dose of 400 mg and this decision is based on the toxicity profile of the drug. This dose is also within the accepted regimen to treat alcoholism. In case of intolerance, lower dose of 200 mg per day is allowed. Patients will take disulfiram in the evening. Patients have to avoid alcohol.

Copper:
Copper supplementation will be given separately from disulfiram (in the morning). Copper nutritional supplement will correspond to 2,5 mg of elementary copper and this is not considered a safety concern 21 . This dose is within the typical copper intake in western countries. WHO has concluded that the fatal oral dose of copper salts is about 200 mg/kg body weight. 21 Thus, the nutritional supplementation of copper in this study is to ensure that copper levels are adequate to keep copper homeostasis with access to generate the Cu-DTC complex also in a patient population where dietary intake may be insufficient.
Concerning chronic toxicity there were no reported problems when the drinking water in in areas over time contained copper at concentration of 8 mg/L. 21 Nevertheless, potential adverse reaction of copper intoxication includes salivation, epigastric pain, nausea, vomiting, diarrhea, renal and hepatic injury.

Explanation for choice of comparator
In the recurrent situation there is no true consensus on what to use. A common approach is to prescribe temozolomide again as long as the progression occurs more than 3-6 months after ended primary treatment, and especially if initial response was observed. However, in cases where progression is seen on ongoing or early after temozolomide treatment it is common practice to prescribe lomustine (also known as CCNU) 5 or PCV (procarbazine, CCNU, vincristine). Lomustine is also the suggested control arm in studies in recurrent glioblastomas due to its established effect in this phase. 22 The downside is hematological toxicities, especially in patients previously treated with temozolomide. 22 Both abovementioned standard treatments have very modest effect in the recurrent setting and new treatments are clearly indicated.
Also MGMT inhibition is seen with disulfiram in preclinical data, and this is predictive of response to alkylating agents.
Consequently, we have chosen any alkylating chemotherapy as the control arm where the intervention groups receive disulfiram and copper supplement given as add-on to chemotherapy.

Justification of choice of trial population
Glioblastoma in the recurrent situation is very treatment resistant and is an immediate threat to patients' survival. No true consensus exists on treatment in the recurrent situation. Clearly, in this situation experimental therapy is justified and especially one with expected low toxicity such as disulfiram hopefully not having serious detrimental effects on quality of survival.

Risk and benefit
It may be a personal sacrifice for patients avoid alcohol intake completely. Further, disulfiram has a well-known risk profile and the major risks of treatment with 400 mg disulfiram daily lie mainly in lack of compliance with respect to alcohol intake and the rare, but potentially serious hepatitis. The risks, also together with chemotherapy in this context, are therefore limited. 20 In addition, there is a risk of neuropathy that is reversible upon drug discontinuation. 20 The potential benefit lies in improved response to treatment. It is the sponsor´s impression that disulfiram treatment compared to regular chemotherapeutic agents has a favorable risk profile and that pursuing clinical studies to elucidate on treatment efficacy is warranted. The sponsor thus concludes that the overall potential benefit clearly outweighs the risks associated with treatment.

Project aim
The project is planned for 6 years and with the following aim: Gaining initial clinical experience of disulfiram efficacy as an active therapy for human glioblastoma, performing the first randomized controlled study with this treatment.
This will be accomplished in a clinical multicenter study carried out at academic hospitals in Sweden and Norway.

Methods
This trial is a multicenter two-armed open labeled 1:1 randomized controlled trial. The study will be conducted in several Swedish and Norwegian oncological units..

Primary end-point
Survival at 6-months analyzed according to intention-to-treat. For evaluation of end-points, see attachment 3.

Secondary end-points
In general, all analyzes will be performed according to intention-to-treat (primary aim) and explored as as-treated analyses. All endpoints will be analyzed with an investigator blinded for the treatment arm. Concerning the primary end-point we do not expect any missing data.

Imaging
MRI according to clinical protocol, including preoperative and postoperative MRI if undergoing repeated resection and with a new MRI scan routinely scheduled every 3-month thereafter up to 24 months post-inclusion. This should be implemented also after documented progression in the disulfiram group if patient continues on disulfiram alone or in combination with another rescue therapy and in the control group as long as tumor targeted therapy is given.

Power estimation
The analysis below is based on data from the BELOB trial (median in lomustine group; 1 month (95 % CI 1-3 months); 13 % PFS at 6 month; median overall survival 8 months (95 % CI 6-11); 30 % overall survival at 12 months). 5 We will use 6 month survival (analyzed at 6 months post-randomization of the last patient included) as primary end-point, but relaxing the alpha value to 0.10 (by convention 0.05) due to the phase II setting. 25 As stated by Khan et al; phase II studies are "preliminary assessment of a new intervention before embarking on a larger and expensive randomized controlled trial", 25 thus slightly increasing the chance of false positive results (due to chance).

Sample size
Based on the assumption that the treatment group will have an improvement in proportion achieving 6-month survival from 60 % to 80 % we need a final sample size of 128 in total (64 in each group; alpha 0.1, power 80 % and two sided test, see below). Considering 10 % attrition the actual included sample need to be 142 with 71 patients included in each group.

Inclusion criteria
The subjects must fulfill all the following inclusion criteria to be eligible for participation in the study, unless otherwise specified: for glioblastoma at the moment of inclusion or during active treatment within the assigned group (i.e. control or disulfiram group). 6. Able to take oral medications. 7. No known allergy to disulfiram or copper. 8. Absolute neutrophil count ≥ 1,500/mcL and platelets ≥ 100,000/mcL 9. Willing to refrain from ingestion of alcoholic beverages while on the study is a criteria to be randomized. However, once randomized alcohol abstinence only affects the group treated with disulfiram, and in this group it includes the entire period and one month after last dosage of disulfiram.

Exclusion criteria
Potential study subjects who meet any of the following criteria are not eligible for participation in the study: 1. Earlier chemotherapy for progression (e.g. "rescue therapy") 2. History of idiopathic seizure disorder, psychosis or schizophrenia.
3. History of uncontrolled hypertension (i.e. systolic BP > 180 mmHg) and a diagnosis of congestive heart failure 4. Received radiotherapy within the 3 months before the diagnosis of progression (see also attachment 4). 5. Addiction to alcohol or drugs. 6. Pregnant and/or breastfeeding. 7. Women of childbearing potential who do not have a negative pregnancy test taken no longer than 14 days prior to randomization. 8. History of active liver disease, including chronic active hepatitis, viral hepatitis (hepatitis B, C and CMV), cholestatic jaundice of any etiology or toxic hepatitis or inadequate hepatic function, defined as baseline ASAT and ALAT > 2.5 X upper institutional limit and/or bilirubin > 2.0 X upper institutional limit. 9. History of Wilson's disease or family member with Wilson's disease (unless excluded as a carrier by genetic test). 10. History of hemochromatosis or family member with hemochromatosis (unless excluded as a carrier by genetic test). 11. Nickel hypersensitivity (disulfiram mobilize nickel causing a brief increase in nickel concentrations before excretion. The initial increase may lead to hepatitis in predisposed patients). 7 12. Need for metronidazole, warfarin and/or theophylline medication (the metabolism may be influenced by disulfiram). 13. Patients who are taking medications metabolized by cytochrome P450 2E1, including chlorzoxazone or halothane and its derivatives (phenytoin, phenobarbital, chlordiazepoxide, imipramine, diazepam, isoniazid, metronidazole, warfarin, amitriptyline within 14 days prior to the first dose of disulfiram. Of note, lorazepam and oxazepam are not affected by the P450 system and are not contraindicated with disulfiram). 14. Unfit for participation for any other reason judged by the including physician.

Informed consent
Patients will be included only after the informed consent is signed (see attachment 6a: Swedish and 6b: Norwegian). The informed consent needs to be dated at time of inclusion, be explicit and written and may at any time be withdrawn by the participant. Patients will be asked to participate during an outpatient visit to the local neurologist or oncologist. Also, the right of the participant to refuse to participate without giving reasons must be respected.
Patients without possibility to provide written, informed consent (e.g. severe cognitive deficit and aphasia) are not to be included via proxies.

Handling of the experimental drug and the nutritional supplement
The experimental therapy consists of disulfiram treatment and nutritional copper supplement. In Norway and Sweden, copper supplement will be delivered to study participants randomized to the experimental arm by the local investigator. Based on the safety profile and long experience with the experimental drug the local investigator prescribes Disulfiram, and the necessary information will be included in the prescription. Patients will collect disulfiram at the pharmacy similar to any other prescription. In Sweden due to a reimbursement arrangement patients will not experience extra expenses ("högkostnadsskyddet"). Norwegian sites will also prescribe the drug from the pharmacy and the batch number of disulfiram will be registered by the pharmacy in a log available also to the local investigator upon request. After agreement with the local hospital pharmacy the cost of the drug will be charged the sponsor directly.

Data Collection
Data will be recorded in web-based case report forms (eCRFs). A data collection manual will be supplied to all centers and each center is responsible to know the content and the routines for all data collection.
All CRFs will be accessible by sponsor to assess data entry, review of drug safety and ultimately, statistical analysis.

Blood samples
Blood samples to be samples are consider clinical routine and taken at routine intervals: Hemoglobin, thrombocytes, leukocytes with differential count, ASAT, ALAT, GT, ALP, bilirubin and INR.

In case of temozolomide treatment
In these cases we plan assessment every 4 weeks to coincide with clinical routine.

In case of lomustine/PCV treatment
There will not be monthly follow up, but at every 6 weeks to follow the clinical routine. We will register the first control at 1 month, then at 3 month, 4 and 6 months and so on. This is chosen to minimize patient burden and may lead to 4 more observations in the temozolomide group than in the lomustine group. Still, the visit planned every third month with MRI is identical.

Delay in treatment
In case of co-morbid events leading to delay or breaks in treatment the assigned treatment is started when appropriated as judged by the local physician. The patient is entered into the study schedule at the nearest time-point to actual treatment with no additional visits planned to correct for this to minimize patient burden. Further entry into study then follows the study schedule.

Compliance
Compliance of disulfiram and copper treatment will be assessed by tablet count at controls in addition to patient self-reporting. If less than 50 % of expected tablets are consumed the patient will be considered non-compliant (in the given period), and if the patient at follow-up visit then wish to withdraw the patient will be regarded as cross-over from last visit (including non-compliant period). This is for descriptive analyses, or for post hoc analyzes, since the pre-defined end-points are analyzed according to intention-to-treat.

Modification in disulfiram treatment before progression or in case of glioblastoma progression
Treatment with disulfiram is continued as long as any tumor targeting treatment is indicated, (i.e. until patient moves into strict palliative phase).
• In case of discontinuation of alkylating chemotherapy before any signs of progression (e.g. due to adverse events specific to chemotherapy including hematological toxicities) treatment with disulfiram is continued either as a single agent or combined with another chemotherapy as long as this is outside of a clinical trial. In fact, the treating oncologist is encouraged to combine further disulfiram treatment with another second line rescue therapy (at the discretion of the treating physician). • If progression is documented, change of chemotherapy may occur. In case of prescription of new cytotoxic treatment (2 nd line rescue therapy), disulfiram should be continued in the group randomized to disulfiram treatment.
In all cases where disulfiram is continued, but with modifications from the original assigned treatment, the patient should be followed with visits, imaging and blood samples according to the protocol.

Cross-over
Before study closure no crossover from the control group to disulfiram is allowed.

Discontinuation/stopping-rules
Thus, treatment with disulfiram is only to be discontinued when: • Unacceptable adverse effect(s) as judged by the treating doctor • No longer indication for tumor targeting therapy • The patient wishes to withdraw from treatment • In case of certain grade 3 and 4 adverse events believed to be caused by disulfiram treatment or put patients on disulfiram treatment to excess risk (see below). • Other Discontinuation criteria for disulfiram at the individual level due to adverse effect(s) • Hepatobiliary adverse effect grade 3 and 4.
• Development of congestive heart failure grade 3 and 4.
• New onset delirium and/or psychosis grade 3 or 4 adverse effect not induced by increasing dose of corticosteroids.

• Other grade 3 or 4 adverse effects where treating physician recommend discontinuation of disulfiram
Reasons for discontinuation of assigned treatment (control group) • Unacceptable adverse effect(s) as judged by the treating doctor • No longer indication for tumor targeting therapy • The patient wishes to withdraw from treatment • Other

Interim analysis and abortion criteria
At 50 % inclusion (71 included patients) an interim analysis will be performed. In this analysis serious adverse events will be explored. A data monitoring scientific committee (DMSC) independent of sponsor and local investigators will be set up to review confidentially the results of this interim analysis (see attachment 7).

Randomization
This is an open-label trial (i.e. no placebo) with alkylating chemotherapy in the group randomized to control and alkylating chemotherapy with add-on of disulfiram in the group randomized to intervention.
Patients will be randomized separately at each study center. Blocks of varying sizes will be used to make prediction of allocation impossible. Randomization will be performed by a web-based randomization system developed and administered by Unit for Applied Clinical Research, Norwegian University of Science and Technology.

Blinding procedures
The trial is open labeled; however a statistician blinded for treatment assignment will perform data analyses. A radiologist blinded for treatment allocation will evaluate radiological endpoints.

Other methods
We will not stratify for more than study center, however important base-line variables like repeated surgery, time since primary treatment, age, functional status, lomustine versus temozolomide, biomarker status from initial surgery if available (IDH and MGMT) will be registered and adjusted for using post-hoc exploratory analyses in case of skewed randomization.

Study schedule
As seen from the study schedule the patients are in the trial follow-up for 24 months. Earlier reason for discontinuation of follow-up includes death or any criteria mentioned in the 3.11.4 section above. Even though the patients return to current clinical practice after 3.11.4 discontinuation, we will register progression and survival since they are main study outcomes.
To minimize patient burden in case of treatment discontinuation we will not continue to measure quality of life. • Alcohol screen relevant if treated with disulfiram • QoL assessed with EQ-5D until progression OR as long as on disulfiram treatment in case of treatment also beyond progression (i.e. together with 2 nd line rescue therapy). • Compliance assessed by disulfiram and copper tablet intake (by tablet count at controls) and patient self-reporting • Routine assessment of adverse events indicated here as long as on active treatment either with alkylating chemotherapy (control group) or disulfiram (intervention group) • Blood samples as indicated here with hematological and liver monitoring relevant as long as on active treatment (Hemoglobin, thrombocytes, leukocytes with differential count, ASAT, ALAT, GT, ALP, bilirubin, INR). The interval for patients treated with lomustine/PCV is 6 weeks, while 4 weeks is clinical routine in the case of temozolomide

General remark on safety of interventional treatment
In the drug information on disulfiram (www.felleskatalogen.no and www.fass.se) it is indicated that severe organic brain disease is a contraindication. This may be true in the treatment of alcohol dependency, but studies on compassionate use in glioblastoma patients describes this as well tolerated. 18 Consequently, there is reason to believe that in the clinical setting without alcohol dependency this is not a significant clinical problem.
Concerning the nutritional supplement with copper this is generally not considered a problem in the dose described here (2,5 mg). 21 Nevertheless, potential adverse reaction of copper intoxication includes salivation, epigastric pain, nausea, vomiting, diarrhea, renal and hepatic injury.
Routine blood samples in all patients every 4-6 weeks (depending on lomustine or temozolomide based treatment; see above) to screen hematological and liver function. This is considered clinical routine.
In addition, due to the well-known profile of disulfiram we will specifically look for 1. Uncontrolled seizures 2. Psychosis or delirium not induced by increasing corticosteroids

Definition adverse event
An adverse event is any untoward medical occurrence in a subject to whom a medicinal product has been administered, including occurrences which are not necessarily caused by or related to that product. The procedure of adverse event reporting is covered in detail by attachment 8.
Due to expected toxicity of the treatments and symptoms related to cancer diagnosis, only grade 3, 4 and 5 (CTCAE) or grade ≥ 2 for infections should be reported.

Toxicity
Adverse events and toxicities must be graded according to the NCI Common Toxicity Criteria for adverse events (NCI-CTCAE) Version 4.0 (http://ctep.cancer.gov/reporting/ctc.html).

A Serious Adverse Event (SAE)
A medical occurrence or effect that at any dose falls in one or more of the following categories: • Results in death • Is life-threatening • Requires hospitalization or prolongation of existing inpatients' hospitalization. Hospitalization refers to a situation whereby an AE is associated with unplanned overnight admission into hospital. • Results in persistent or significant disability or incapacity • Is a medically significant adverse event It is the responsibility of the local investigator to collect all the above-mentioned adverse events. Adverse events unequivocally due to standard of care treatment (see attachment 8) will not be reported as adverse events. Also, for definitions of categories of SAEs, also see attachment 8.
Still, events based on the knowledge of the disease in question and expected clinical course some events otherwise classified as serious are not considered/classified as SAE in this study. The following is a list of SAEs, which do not require to be reported as SAEs: • Events based on the knowledge of the disease in question and expected clinical course (table 2). • If a patient is admitted to hospital with a documented cancer related problem then this will not be reported as a SAE (table 2). In every reportable case of SAE the investigator is responsible for ensuring that reporting to the Sponsor occurs no later than 24 hours after any of the site staff become aware of the event. Initial reports should be followed as soon as possible by detailed written reports describing the nature of the event and clinical consequences.

Serious Adverse Reaction (SAR) and Suspected Unexpected Serious Adverse Reactions (SUSAR)
A SAR is an SAE that may be related to trial treatment. The assessment of "relatedness" is primarily the responsibility of the Principal Investigator at site or agreed designee (table 3).
SUSAR are reactions where disulfiram (or copper supplement) may be suspected as the reason for an unexpected SAR. Both SAR and SUSAR are considered more serious and have to be reported to the sponsor immediately (within 24 hours). The sponsor is responsible to report to the respective national authorities within their time limits.
SAEs that will be considered related (i.e. SAR or SUSAR) include any SAE that is documented as possibly, probably or definitely related to protocol treatment. Table 3: The assessment of relatedness is made using the following:

Description
There is no evidence of any causal relationship. There is some evidence to suggest a causal relationship (e, g. the event occurs within a reasonable time after administration of the trial medication). However the influence of other factors may have contributed to the event (e.g. the patient's clinical condition, other concomitant treatments). There is evidence to suggest a causal relationship and the influence of other factors is unlikely.
There is clear evidence to suggest a causal relationship and other possible contributing factors can be ruled out.

SAR and SUSAR are considered more serious because of the relatedness and have to be reported to the sponsor immediately without delay (within 24 hours). The sponsor is responsible to report to the respective national authorities within their time limits.
For an unexpected and serious adverse event that was lethal or life threatening, immediate and no later than 7 days after being notified the sponsor has to report to authorities in all involved EEA countries. Relevant follow-up information concerning further clinical progress has to be given no later than 8 days after the initial report. Also, sponsor is obliged to send immediately and no later than 15 days after being notified of an unexpected and serious adverse event that is not lethal or life threatening to authorities in all involved EEA countries. In addition, sponsor is obliged to report back to local investigators of suspected adverse events that are serious and unexpected (i.e. SUSAR).

Special considerations for women of childbearing potential
As mentioned in exclusion criteria 6 and 7 pregnant patients or patients that are breastfeeding cannot be included in this trial. As stated, women of childbearing potential must also provide a negative pregnancy test before enrollment. Further, women of childbearing potential have to use a highly effective birth control method as defined by the European Heads of Medicines Agencies, Clinical Trial Facilitation Group document concerning "Recommendations related to contraception and pregnancy testing in clinical trials", paragraph 4.1, page 9 (http://www.hma.eu/fileadmin/dateien/Human_Medicines/01-About_HMA/Working_Groups/CTFG/2014_09_HMA_CTFG_Contraception.pdf).

Trial conduct
This study will be conducted in compliance with the protocol and according to Good Clinical Practice guidelines and applicable regulatory standards. Specifically, the Swedish and Norwegian ethical committees and the national medicine agencies need to approve the study. Further, the study will be conducted in accordance with the latest Declaration of Helsinki. 26

Case report forms
All data will be entered in the eCRF at the local study site. The Unit for Applied Clinical Research at the Norwegian University for Science and Technology (Trondheim, Norway) will manage the eCRF.

Patient security
All patient information is handled using patient ID that is not logically connected to any personal data. It is possible to "decode" back to personal data using a name list stored securely at the respective study sites. In the eCRF patients will be identified with a study participation number.

Specification of source documents
The following documents are source documents -Patient hospital records -Radiology reports -MRI scans -Lab reports -Chemotherapy registers -Radiotherapy registers -The quality of life questionnaires

Archiving
All data will be stored at the participating sites for 15 years. In addition the electronic data will be kept during this time.

Data management
The CRFs in this trial are implemented electronically (i.e. eCRF) using a dedicated software developed at the Unit for Applied Clinical Research at the Norwegian University for Science and Technology (NTNU). This solution is at present (December 2015) used by more than 120 ongoing studies. All data entered into the eCRF are stored at two separate servers at the NTNU. These servers are protected by the general firewall at the university and then by another firewall dedicated to protect clinical research data. The server that stores the data is not the same server as the one in contact with the Internet. There are several further safety measures not to be exposed. There are backups taken every 24 hour throughout the years. The responsibility for this system lies with the Unit for Applied Clinical Research at NTNU, Trondheim, Norway.

Analysis
At time of analysis, data files will be extracted from the database into statistical packages to be analyzed.

Monitoring
All source data must be accessible for auditing and monitoring. Monitor will maintain patient confidentiality. For details, see attachment 9.

Inspection
Authorities have the right to perform inspections as well as the monitor. The inspector will have access to all relevant source data and study material. The local responsible investigator will answer any questions arising. However, the sponsor/principal investigator will be available for questions/dialogue in the immediate period following such inspections as well.
All involved parties will keep the patient data strictly confidential.

Premature Termination of the Study
The sponsor may terminate the study prematurely for any reasonable cause. The Ethics committees and competent authorities will then be informed promptly. Conditions that may warrant termination include, but are not limited to: • The discovery of an unexpected, significant, or unacceptable risk to the patients enrolled in the study. • If competent authorities obtain information that raises doubts about the safety or scientific validity of the study, the competent authorities can suspend or prohibit the study.

Insurance
Patients will be insured according to national requirements.
Norway: www.laf.no (Legemiddelansvarsforeningen, LAF = the Drug Liability Association), see attachment 10. The sponsor has made a specific insurance that covers all Norwegian study participants.

Regulatory requirements
The respective ethical committees at the study sites need to approve the study. In addition, the medical product agencies (the Swedish Medical Products Agency (MPA) and the Norwegian Medicines Agency (NMA)), have to authorize the study. Enrolment of patients will not start until approval has been received from both the Ethics Committee(s) and Competent authorities.
It may be desirable to make changes in the study at a later stage. In that case, the medical product agencies and ethical committees must approve significant changes in the protocol before they are implemented. We will use the form that is common across the EU (Substantial Amendment Notification Form) in reporting to the medical product agencies. A substantial change, is for example, change of the main purpose, primary or secondary variables, method to measure the primary endpoint, change of investigational medicinal products or dosage.

Cost
Due to the cheap treatment and the small change in follow-up routines the costs will be taken through regular clinical practice or from research grants provided by sponsor.
Administration of project, data monitoring, randomization process, data management, statistics and radiology services will be covered by sponsor through research grants. The project has a total budget of approximately 4 Mill SEK.

Estimated progress
Year